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STUDIES  ON  HOG  CHOLERA 


CHARLES    McCLINTOCK 

CHARLES  H.  BOXMEYER 

J.  J.  SIFFER 

(From  the  Department  of  Experimental  Medicine,  Parke,  Davis  &  Co.,  Detroit) 


Reprinted  from 
The  Journal  of  Infectious  Diseases,  Vol.  II,  No.  2,  March  i,  1905,  pp.  351-374 


CHICAGO 


irv  i 


STUDIES  ON  HOG  CHOLERA.* 

Charles  T.  McClintock,  M.D.,  Charles  H.  Boxmeyer,  A.B., 

AND   J.   J.    SlEFER,   M.D. 
(From  the  Department  of  Experimental  Medicine,  Parke,  Davis  <&  Co.,  Detroit.) 


THE  VALUE  OF  AGGLUTINATION  FOR  THE  DIAGNOSIS  OF 

HOG  CHOLERA. 

Charles  T.  McClintock,  Charles  H.  Boxmeyer  and  J.  J.  Sipper. 

The  close  resemblance  of  the  hog  cholera  to  the  typhoid  bacil- 
lus, and  the  somewhat  similar  symptoms  and  course  of  the  diseases 
caused  by  them,  suggested  that  agglutination  might  prove  as 
valuable  a  method  of  diagnosis  for  hog  cholera  as  it  has  proven 
for  typhoid.  The  first  application  of  the  agglutination  test  for 
tliis  purpose  was  made  by  Dinwiddle,^  of  the  Arkansas  Experi- 
ment Station.  The  few  cases  he  tested  all  gave  a  negative  reac- 
tion. No  hog  cholera  bacilli  were  isolated  from  the  outbreak 
which  furnished  this  material.  Dawson,"  Smith,^  and  others  have 
made  experiments  with  the  blood  of  vaccinated  rabbits  which 
show  the  reaction  often  in  high  dilutions.  We  are  unaware  of 
any  other  experiments  than  those  of  Dinwiddle  with  the  blood  of 
diseased  hogs. 

The  material  with  which  we  worked  was  obtained  from  the 

following  seven  epizootics: 

1.  Hastings,  Mich. —  The  disease  first  appeared  in  slaughter-house  hogs 
fed  on  refuse.  It  was  communicated  by  them  to  hogs  on  an  adjoining  lot  sepa- 
rated only  by  a  wire  fence.  From  there  it  spread  to  neighboring  farms. 
Practically  all  the  animals  exposed  contracted  the  disease.  The  mortality, 
including  animals  of  all  ages,  was  50  to  60  per  cent.  The  animals  that  sur- 
vived apparently  thrived.  Autopsy  showed  considerable  variation  in  the 
pathological  appearances  of  the  individual  animals  examined.  Practically  all 
the  lesions  described  for  hog  cholera  were  found.     The  intestinal  changes  as 

*  Received  for  publication  January  14,  1905. 
1  Jour.  Covip.  Med.  and  Vet.  Arch.,  1900,  21,  p.  528. 
^Neio  York  Med.  Jour.,  February  20,  1897,  65,  p.  253. 
3  Jour.  Med.  Research,  1903,  91,  p.  270. 

351 


a^i^a 


352  McClintock,  Boxmeyer,  and  Siffer 

a  rule  were  quite  marked.  A  marked  hyperemia  only;  small  deep  ulcers, 
large  diphtheric  patches,  areas  denuded  of  epithelium,  a  diphtheric  mem- 
brane covering  the  entire  cecum  and  colon,  raised  button  ulcers,  fungoid 
growths,  and  follicular  abscesses  were  observed.  Pneumonitis,  pleuritis,  and 
pericarditis  were  frequently  found.  Edematous  and  hemorrhagic  lymph 
glands  were  practically  constantly  present.  The  spleen  was  usually  small 
and  the  kidneys  showed  a  pale  cortex,  frequently  with  hemorrhagic  spots. 
These  latter  were  variations  from  the  usual  description  of  hog  cholera.  Swine 
plague  bacilli  were  obtained  from  the  hogs  showing  lung  lesions. 

2.  St.  Johns,  Mich. — Hog  cholera  had  been  prevalent  in  this  locality  for 
several  years.  From  July  to  November  two  townships  had  lost  about  1,000 
head.  The  loss  was  about  60  per  cent.  The  presence  of  a  severe  diarrhea 
characterized  this  outbreak.  Autopsy  on  three  animals  showed  lesions  simi- 
lar to  those  seen  in  the  Hastings  outbreak.  Lung  and  intestinal  lesions  were 
both  present. 

3.  Chatham,  Ont. — Here  also  the  disease  had  existed  for  several  seasons. 
During  the  summer  severe  losses  had  occurred.  In  December,  at  the  time 
we  visited  the  locality,  the  disease  had  ceased  to  spread.  The  mortality 
exceeded  50  per  cent.  On  account  of  the  severe  cold  weather  and  the 
Dominion  quarantine  laws,  but  two  autopsies  were  made.  Lung  lesions  were 
most  marked.  The  Canadian  inspectoi's  spoke  of  intestinal  and  foot  lesions 
as  occurring. 

4.  Belle  Plaine,  la. — George  W.  Dunphy,  D.  V.  S.,  described  this  out- 
break as  follows:  "This  is  a  very  virulent  form  of  a  mixed  hog  cholera  and 
swine  plague,  destroying  more  than  90  per  cent  of  each  herd  in  which  it 
appears.  In  the  first  autopsies  only  swine  plague  lesions  were  found,  but 
later  I  discovered  well  marked  hog  cholera  lesions.  In  the  first  postmortems 
the  stomach  and  intestines  were  practically  normal.  The  lungs  were  com- 
pletely hepatized.  On  section  pus  exuded  from  the  bronchi  and  a  bloody 
fluid  from  the  lung  tissue.  The  liver  and  spleen  were  normal  in  appearance. 
In  the  later  postmortems  I  found  extensive  congestion,  ecchymoses  and  thick- 
ening of  the  intestinal  mucosa,  also  the  characteristic  projecting  button 
ulcers  of  hog  cholera.  In  the  very  acute  cases  there  were  extensive  intes- 
tinal hemorrhages,  and  large  blood  clots  were  found  among  the  intestinal 
contents." 

5.  Lincoln,  Neb. — This  outbreak  occurred  at  the  Nebraska  Experiment 
Station.  The  disease  was  introduced  by  a  pig  purchased  at  the  State  Pair. 
A  heavy  loss  occurred.  Dr.  Dunphy  described  the  lesions  as  similar  to  those 
found  in  the  Belle  Plaine,  la.,  epizootic.  Both  intestinal  and  lung  lesions 
occurred. 

6.  TeJconsha,  Mich. — Two  hogs  shipped  by  Dr.  Dunphy  to  the  laboratory 
were  killed  and  examined.  Both  showed  follicular  abscesses  in  the  large 
intestines,  and  hemorrhagic  lymph  glands.  The  other  organs  were  appar- 
ently normal.  These  were  convalescent  cases.  Hog  cholera  bacilli  were 
not  found. 

7.  State  Insane  Asylum,  Pontiac,  Mich. — This  is  the  outbreak  which  is 
described  in  the  succeeding  section  on  "The  Production  of  Hog  Cholera  in 
Swine  by  Inoculation  with  Bacteria-free  Filtrates,"  q.  v. 


Studies  on  Hog  Choleea  353 

Bacteriological  examinations  were  made  of  a  number  of  ani- 
mals, as  follows: 

Animals 

Tekonsha,  Mich.       ------  2 

Hastings,  Mich.     ------  9 

St.  Johns,  Mich.        ------  4 

Pontiac,  Mich.       ------  4 

Belle  Plaine,  la.         ------  2 

Lincoln,  Neb.         -        -        -        -        -        -  1 

Chatham,  Ont.  -------  1 

From  viscera  from  Belle  Plaine,  la.,  and  Lincoln,  Neb.,  hog 
cholera  bacilli  were  isolated  which  agreed  quite  closely  with  each 
other  in  cultural  and  agglutinative  characters.  These  bacilli  were 
but  moderately  virulent  for  experimental  animals. 

From  the  Pontiac  outbreak  we  isolated  an  organism  of  the  hog 
cholera  group  apparently  identical  in  morphological,  cultural,  and 
agglutinative  characters  with  the  bacillus  /3  of  swine  dysentery 
of  Theobald  Smith, 

In  no  other  instance  were  we  able  to  isolate  true  hog  cholera 
bacilli,  although  a  number  of  organisms  quite  closely  related  were 
found. 

TECHNIQUE. 

Suspensions  of  agar  cultures  16  to  24  hours  old  in  salt  solution  saturated 
with  tri.-  chlor.-  tertiary-  butyl-alcohol  (chloretone)  were  employed  through- 
out this  work.  Dilutions  of  the  serum  to  be  tested  were  made  in  physiological 
salt  solution,  and  equal  parts  of  the  serum  dilution  and  the  bacterial  suspen- 
sion mixed  in  small  test  tubes  (8x80 mm.)  The  tubes  were  then  placed  in  the 
incubator  at  37°  C.  They  were  examined  first  at  the  end  of  two  hours  and 
again  after  16  or  20  hours.  The  highest  dilutions  in  which  plain  floccules 
showed  were  taken  as  the  limit.  Although  the  bacterial  suspension  remained 
fit  for  use  for  many  weeks,  fresh  suspensions  were  usually  made  for  each  test. 
Control  tests  with  young  broth  cultures  and  the  microscope  were  made  from 
time  to  time.  In  a  number  of  instances  dried  blood  was  used;  this  we  found 
reacted  fully  as  well  as  the  fresh  serum. 

THE   PRESENCE   OF  AGGLUTININS   FOR   HOG  CHOLEEA  BACILLI    IN   THE 
BLOOD   OF  NORMAL  HOGS. 

The  blood  of  normal  hogs  usually  agglutinates  hog  cholera 
bacilli  in  fairly  high  dilutions.  For  our  purpose  it  was  necessary 
to  determine  the  'normal  range.  Of-  22  specimens  of  normal  hog 
blood  examined  3  agglutinated  hog  cholera  bacilli  in  dilutions  of 


354 


McClintock,  Boxmeyee,  and  Siffee 


1-200,*  11  in  dilutions  of  less  than  100;  the  remainder  between 
these  limits.  (  The  smaller  amount  of  agglutinin  is  usually  found 
in  the  blood  of  the  younger  hogs.)  The  blood  of  normal  hogs 
agglutinates  not  only  the  hog  cholera  bacillus  but  also  others: 
swine  plague,  colon  and  paracolon  bacilli  in  dilutions  of  about  the 
same  strength  as  the  hog  cholera  organism. -j- 

From  a  consideration  of  these  data  it  was  concluded  that  an 
agglutination  in  a  dilution  of  less  than  1-300  is  of  no  value  in 
the  diagnosis  of  an  infection  by  hog  cholera  bacilli. 

AGGLUTINATIONS  WITH  THE  BLOOD  OF  HOGS  SICK  WITH 
OE  DEAD  OF  HOG  CHOLEEA. 

The  blood  of  diseased  hogs  in  the  outbreaks  studied  reacted 
only  occasionally  with  hog  cholera  bacilli  in  dilutions  of  1-300. 
The  following  table  gives  the  results  of  the  examination  of  the 
blood  of  57  different  hogs  from  the  several  outbreaks: 

TABLE  I. 

The  Agglutinatiox  of  Hog  Cholera  Bacilli  by  The  Blood  of  Diseased  Swine. 


Source  of 
Blood 

No.  Animals 

from  which 

Blood  was 

Tested 

Eeaction  and 
Dilution 

Hos  Cholera 
Bacillus 
Isolated 

Hastings 

7 
3 
2 

7 
3 

\t 

\    5 

n 

1 

-  200 

-  300 

-  250 

-  300 

-  300 
+2000 
+1000 
+  600 

-  300 

-  50 
+  800 

-  300 

no 

St.  Johns 

no 

Takonsha 

Belle  Plaine 

Herd  1 

no 

Herd  2 

Herd  3 

yes 

Lincoln  

Pontiac 

Chatham 

yes 

yes 
no 

In  those  instances  where  a  hog  cholera  bacillus  was  isolated 
from  an  outbreak,  the  agglutinating  power  of  the  blood  of  the 
animals  from  this  locality  was  tested  against  this  particular 
bacillus.      In  those  outbreaks  from  which  no  hog  cholera  bacillus 

*That  the  presence  of  this  relatively  large  quantity  of  agglutinin  cannot  be  explained 
by  the  animals  having  had  a  previous  attack  of  the  disease  is  shown  by  their  succumbing 
to  hog  cholera  when  subsequently  exposed. 

t  The  Pontiac  bacillus  was  never  agglutinated  by  normal  hog  blood  in  any  of  the  trials 
made  in  dilutions  of  1-50. 


Studies  on  Hog  Cholera  355 

was  isolated  the  agglutinative  power  of  the  blood  was  tested  upon 
several  strains  of  hog  cholera  bacilli. 

Table  I  shows  plainly  that  agglutination  is  of  no  value 
for  the  diagnosis  of  hog  cholera,  as  the  disease  is  at  present 
defined.  The  presence  of  a  positive  reaction,  if  obtained  in  dilu- 
tions of  1-300  or  over,  does,  however,  indicate  an  infection  w^ith 
hos:  cholera  bacilli. 


II. 

THE  PKODUCTION  OF  AGGLUTININS  FOR  HOG  CHOLERA 
BACILLI   IN  SWINE. 

Charles  T.  McClintock,  Charles  H.  Boxmeyer  and  J.  J.  Sipper. 
AGGLUTINATION   AFTEE    VACCINATION    WITH    HOG   CHOLEEA    VACCINE. 

The  vaccine  used  in  these  experiments  was  prepared  by  grow- 
ing 11  different  strains  of  hog  cholera  and  one  of  swine  plague 
bacilli  upon  agar  in  quart  whiskey  flasks.  After  incubating  at 
37°  C  for  60  hours  the  growth  was  loosened  from  the  agar  sur- 
faces with  a  bent  glass  rod  and  washed  off  with  a  .75  per  cent  salt 
solution  saturated  with  tri.-  chlor.-  tertiary-  butyl-alcohol.  The 
suspension  was  then  strained  through  India  mull  to  remove 
particles  of  agar  and  cotton,  the  several  strains  mixed,  and  heated 
to  50°  C  for  30  minutes.  The  vaccinations  were  practically 
always  made  intraperitoneally.  The  animal  was  bled  seven  or 
eight  days  afterwards  from  the  tail  or  ear. 

Most  of  the  animals  reacted  promptly  to  the  injection  pro- 
ducing relatively  large  quantities  of  agglutinin.  A  series  of  four 
hogs  was  vaccinated  with  25  c.c.  of  hog  cholera  vaccine  and  the 
agglutinating  power  of  their  blood  tested  each  day.  The  amount 
of  agglutinin  in  the  blood  began  slowly  to  ris6  on  the  second  and 
third  day,  then  it  rose  rapidly  till  the  sixth  day  when  it  attained  its 
maximum.  It  stayed  at  this  point  for  about  three  days.  During 
the  succeeding  week  it  dropped  either  suddenly  or  gradually  to  a 
relatively  constant  level  at  which  it  remained  for  some  time.  A 
second  vaccination  21  days  after  the  first  raised  the  level  slightly, 
but  it  never  attained  its  former  maximum  height.     This  is  not 


356 


McClintock,  Boxmeyee,  and  Siffee 


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true,  however,  in  all  cases,  the  blood 
of  some  hogs  showing  the  highest  ag- 
glutinating power  after  repeated  injec- 
tions. 

Twenty-six  vaccinated  hogs,  bled 
seven  days  after  injection,  showed  the 
following  maxima: 

3  hogs 75,000  to  100,000 

1  hog  50,000  to    75,000 

3  hogs 20,000  to    50,000 

6  hogs 10,000  to    20,000 

7  hogs 5,000  to    10,000 

6  hogs 1,000  to     5,000 

The  quantity  of  agglutinin  produced 
varied  somewhat  with  the  lot  of  the  vac- 
cine used.  The  size  of  the  dose,  how- 
ever, appears  to  have  had  no  influence 
upon  the  amount  of  agglutinin  produced, 
this  depending  apparently  upon  an  idio- 
syncrasy of  the  animal. 

TABLE  II. 

EELATION  OF  AMOUNT  OF  VACCINE  TO  HEIGHT  OF 
AGGLUTINATION. 


OOOOOOOOc 


O   O  lO   M   rl 


Agglutination 

No.  of 

Amount  of 

Lots  of 

Limit  Seven 

Hogs 

Vaccine 

Vaccine 

Days  after 
Injection 

25  C.C. 

.05183 

50,000 

25    " 

.05183 

10,000 

25    " 

160  H 

5,000 

25    " 

IGO  H 

2,000 

20    " 

.05072  B 

1,200 

20    " 

.05072  B 

300 

12    " 

160  H 

10,000 

12    " 

160  H 

20,000 

12    " 

160  H 

7,500 

10    " 

160  H 

15,000 

10    " 

160  H 

5,000 

10    " 

.05072  B 

1,000 

10    " 

.05072  B 

300 

5    " 

160  H 

10,000 

5    " 

160  H 

5,000 

Agglutinative  value 


Studies  on  Hog  Cholera 


357 


In  a  few  cases  agglutinins  had  not  appeared  in  the  blood  seven 
days  after  vaccination. 

Hogs  inoculated  with  mixed  hog  cholera  and  swine  plague 
vaccine  show  in  their  blood  an  agglutinin  for  the  swine  plague 
as  well  as  the  hog  cholera  bacillus. 

Two  hogs  with  an  agglutinating  limit  before  treatment  of  50 
and  75  for  swine  plague  bacilli  and  100  and  150  respectively  for 
hog  cholera  bacilli,  showed  seven  days  after  vaccination  with 
mixed  vaccine  an  agglutination  of  1-500  and  1-10,000  for  swine 
plague,  and  1-15,000  and  1-50,000  for  hog  cholera  bacilli. 

AGGLUTINATING    POWEE    OF    THE    BLOOD    AFTEE    INOCULATION    WITH 
LIVE    HOG    CHOLEEA    BACILLI. 

Hog  No.  103,  weighing  75  pounds,  received  into  the  ear  vein 
six  c.c.  of  an  18-hour  culture  of  a  hog  cholera  bacillus  obtained 
from  the  Lincoln  (Nebraska)  outbreak,  but  a  few  weeks  after  its 
isolation.  The  animal  was  bled  to  death  on  the  13th  clay.  The 
following  curve  shows  the  agglutinating  action  of  its  serum  upon 
various  strains  of  hog  cholera  bacilli: 


bcQ 


a 

0 

oZ 

> 

0 

Z; 

o 

"3 

pq 

oZ 

3 
> 

cd 
< 

75.000 

^ 

50,000 
20,000 

/ 

^^ 

10,000 

r 

1 

1 

/ 

200 
50 

^ 

y 

A  hog  injected  subcutaneously  with  five  c.c.  of  the  Pontiac 
bacillus  agglutinated  seven  weeks  afterward  the  injected  bacillus 
in  a  dilution  of  1-800,  but  did  not  agglutinate  the  Arkansas 
bacillus  in  a  dilution  of  1-50. 


358 


McClintock,  Boxmeyee,  and  Siffee 


The  following  curve  of  agglutination  by  the  serum  of  a  rabbit 
inoculated  with  the  Pontiac  organism  shows  graphically  the 
agglutination  relations  of  the  hog  cholera  and  swine  dysentery 
bacilli : 


Pierce 

Pontiac 

Sprague 

Arkansas 

400 

?0() 

100 

\^ 

50 

-^ 

MAY    HOG    CHOLEEA    BACILLI     BE    PEESENT     IN    THE    ORGANISM    AND 
THE    AGGLUTINATION    REACTION    REMAIN    NEGATIVE? 

Case  l.—  Hog.  No.  106  was  fed  viscera  of  hogs  in  which  the  Arkansas  hog 
cholera  bacillus  was  present.  He  died  seven  days  afterward.  From  the 
spleen,  liver,  and  mesenteric  glands  pure  cultures  of  the  same  hog  cholera 
bacillus  were  obtained.  The  blood  serum  of  this  animal  gave  no  reaction 
either  with  the  bacillus  isolated  or  with  the  original  culture.  This  observa- 
tion was  confirmed  by  a  repetition  of  the  test. 

Case  2. —  Hog  No.  105  received  intravenously  six  c.c.  of  the  juice  of  the 
organs  of  hog  48,  filtered  through  paper.  He  was  dead  on  the  13th  day. 
Autopsy  show^ed  typical  swine  plague  lesions.  Cultures  from  the  various 
organs  gave  a  mixture  of  the  swine  plague  and  hog  cholera  bacillus.* 

The  blood  serum  gave  no  agglutination  in  dilutions  of  1-300 
with  the  hog  cholera  bacillus  isolated. 

The  blood  serum  of  two  hogs  from  the  Pontiac  outbreak,  from 
which  hog  cholera  bacilli  were  isolated,  gave  no  agglutination 
either  with  the  Pontiac  organism  or  strains  of  ordinary  hog 
cholera  bacilli  in  dilutions  of  1  to  50. 


*The  Arkansas  bacillus,  which  was  kindly  sent  us  by  Dr.  Theobald  Smith,  is  admira- 
bly adapted  for  experimental  work  on  account  of  the  readiness  with  which  it  may  be 
identified.  It  differs  markedly  in  the  appearance  of  its  colonies  from  all  other  strains  we 
have  studied.  On  broth  it  forms  a  thin,  white,  brittle  membrane.  On  agar  it  forms 
characteristic  dry,  slightly  wrinkled  colonies,  readily  detached  in  one  piece  from  the  agar 
surface.  It  is  difficult  to  break  up  the  growth  and  make  up  a  homogeneous  suspension. 
Its  action  upon  various  culture  media  and  toward  agglutinating  serum  was  almost  identi- 
cal with  that  of  the  other  strains  in  our  possession.  The  peculiarities  were  retained 
through  repeated  animal  passages. 


Studies  upon  Hog  Choleea  359 

SUMMARY. 

1.  The  serum  of  normal  hogs  agglutinates  strains  of  ordinary 
hog  cholera  bacilli  in  dilutions  occasionally  as  high  as  1  to  250. 
For  this  reason  we  consider  a  reaction  in  a  dilution  of  less  than 
1  to  300  without  diagnostic  value. 

2.  The  bacillus  of  swine  dysentery  is  not  agglutinated  by 
normal  blood  in  such  high  dilutions. 

3.  Asra'lutination  is  of  no  value  for  the  diagnosis  of  hog  chol- 
era,  as  the  disease  is  at  present  defined. 

4.  The  presence  of  a  positive  reaction  does,  however,  indicate 
an  infection  with  hog  cholera  bacilli. 

5.  There  are  occasional  instances  of  both  natural  and  artificial 
infection  in  which  no  increase  of  the  agglutinins  for  hog  cholera 
over  those  normally  present  can  be  demonstrated. 

6.  The  maximum  amount  of  agglutinin  develops  in  a  hog's 
blood  within  six  or  seven  days  after  a  single  inoculation  with 
hog  cholera  vaccine. 

7.  Hogs  react  to  intraperitoneal  injections  of  hog  cholera 
vaccines,  usually  with  the  production  of  large  quantities  of 
agglutinins,  the  amount  of  the  vaccine  bearing  no  relation  to  the 
amount  of  agglutinin  produced. 


III. 

THE  PRODUCTION  OF  HOG  CHOLERA  IN  SWINE  BY  INOCU- 
LATION WITH  BACTERIA-FREE  FILTRATES. 

Charles  H. Boxmeyer. 

Repeated  failures,  after  thorough  and  prolonged  search,  to 
isolate  hog  cholera  bacilli  from  the  carcasses  of  hogs  which  had 
succumbed  to  the  disease  in  certain  Michigan  outbreaks,  the  fail- 
ure of  the  blood  serum  of  any  of  these  animals  to  agglutinate  any 
of  several  hog  cholera  strains,  together  with  the  inability  to  pro- 
duce the  characteristic  lesions  of  hog  cholera  by  the  feeding  or 
injection  of  hog  cholera  bacilli,  led  to  a  search  for  some  other 
agent  as  the  etiological  factor  of  these  outbreaks. 

Opportunely,  the  Bureau  of  Animal   Industry'  at  this  time 

'  DeSchweinetz  and  Dorset,  Bureau  of  Animal  Industry,  1£03,  Circular  41. 


360  McClintock,  Boxmeyee,  and  Siffee 

published  a  preliminary  report  upon  "A  Form  of  Hog  Cholera 
Not  Caused  by  the  Hog  Cholera  Bacillus,"  in  which  they  stated 
that  they  were  able  to  transmit  the  disease  to  healthy  animals 
"by  the  subcutaneous  inoculation  of  certain  body  fluids,  these 
fluids  being  always  proved  by  careful  bacteriological  examination, 
by  filtration  through  the  finest  porcelain  filters,  and  by  the  inocu- 
lation of  guinea  pigs  and  rabbits,  to  be  free  from  hog  cholera  and 
swine  plague  bacilli." 

Experiments  were  immediately  begun  along  this  line,  as  the 
confirmation  of  this  work  would  explain  the  failure  of  all  attempts 
to  immunize  hogs  by  means  of  vaccines  or  sera  against  hog 
cholera,  and  point  out  the  way  to  a  possibly  successful  method  of 
combating  this  disease. 

experiments  with  filtered  body  fluids. 

The  infectious  material  employed  was  derived  from  an  epizootic 
of  hog  cholera  that  occurred  at  the  State  Insane  Asylum  at  Pon- 
tiac,  Michigan,  in  May,  1904,  At  the  time  of  investigation  the 
disease  had  been  current  for  about  six  weeks.  The  herd  had 
originally  contained  90  head  —  Berkshires,  Chester- whites,  and 
crosses  —  weighing  from  100  to  200  pounds.  Fifty  deaths  had 
occurred,  and  practically  all  the  remaining  hogs  showed  evidence 
of  a  more  or  less  advanced  stage  of  the  disease.  The  symptoms 
shown  were  fever,  reddening  of  the  ears  and  skin  covering  under 
surface  of  the  body,  partial  paralysis  of  the  posterior  extremities, 
and  eyelids  swollen  and  glued  together.  Some  animals  exhibited 
a  marked  constipation,  while  others  showed  a  severe  diarrhea. 
The  superintendent  kindly  permitted  us  to  kill  two  of  the  sickest 
animals  for  postmortem  examination.  Autopsy  showed  the  fol- 
lowing : 

Hog  No.  1. —  Chester-white,  male,  weight  100  pounds.  Had  shown  dis- 
ease symptoms  for  about  one  month.  Condition  fair.  Weakness  of  the  hind 
legs.  Killed  and  examined.  Skin  shows  slight  flush.  Inguinal  glands  not 
enlarged.  Spleen  hyperplastic.  Liver  pale,  of  a  yellowish  color.  Kidneys 
with  pale  cortex  dotted  with  minute  petechiae.  Mesenteric  glands  edema- 
tous and  hemorrhagic.  Cecum  and  colon  filled  with  hard,  impacted  feces. 
Mucosa  dark  colored.  Two  indurated  ulcers  about  15  m.m.  in  diameter  at 
the  base  of  the  ileocecal  valve.  Echinorhynchi  and  ascarides  present.  Small 
intestines  little  changed.  Apices  of  both  lungs  hepatized,  with  pleural  adhe- 
sions at  these  points.     Visceral  pericardium  petechiated. 


Studies  on  Hog  Cholera  361 

Hog  No.  2. —  Berkshire,  weight  125  pounds.  Marked  weakness  in  both 
hind  and  fore  legs.  Condition  fair.  Killed  and  examined.  Inguinal  lymph 
glands  not  enlarged.  Spleen  slightly  enlarged  and  covered  with  fleshy  villi. 
Kidneys  with  pale  cortex  sparsely  petechiated.  Liver  pale.  Intestinal 
mucosa  shows  no  marked  change,  and  intestines  are  filled  with  a  large  quan- 
tity of  semi-fluid  feces.  Lungs  show  fairly  large  sub-pleural  ecchymoses. 
The  posterior  lobe  of  left  lung  is  congested  and  shows  beginning  of  hepatiza- 
tion.    Lymph  glands  swollen  and  slightly  hemorrhagic. 

Cultures  from  the  various  viscera  gave  a  variety  of  organisms,  from 
which  we  were  able  to  isolate  a  hog  cholera-like  bacillus,  identical  in  mor- 
phological, cultural,  and  agglutinating  characters  with  the  bacillus  of  swine 
dysentery  /3  of  Theobald  Smith.  One-half  c.c.  of  a  24-hour  culture  of  this 
bacillus  injected  subcutaneously  into  a  rabbit  produced  death  in  six  days. 
Postmortem  showed  the  focal  necroses  of  the  liver  characteristic  of  an  infec- 
tion by  members  of  the  hog  cholera  group.  The  blood  serum  of  this  rabbit 
agglutinated  the  injected  bacillus  and  the  swine  dysentery  bacillus  ^3  of 
Smith  in  a  dilution  of  1-800,  but  produced  no  agglutination  in  dilutions 
of  1-50  with  the  ordinary  strains  of  hog  cholera  bacilli.  Five  c.c.  of  a  16-hour 
broth  culture  of  this  bacillus  in  the  fourth  generation,  and  four  days  after 
isolation,  produced  no  other  effect  than  a  considerable  local  abscess  when 
injected  subcutaneously  into  a  20-pound  hog  during  the  30  days  he  was  under 
observation.     The  animal  gained  rapidly  in  weight  during  this  period. 

Portions  of  the  spleen  and  liver  of  the  above  hogs  were  brought  to  the 
laboratory  and  fed  to  hog  No.  114,  a  Chester-white  pig,  weighing  20  pounds. 
He  was  chloroformed  on  the  29th  day  after  infection,  being  moribund.  The 
blood  was  collected  aseptically  from  the  carotid  artery. 

Postmortem:  Great  emaciation.  Ears  and  snout  blackened  and  necrotic. 
Feces  thin,  yellow,  fluid.  Inguinal  glands  enlarged,  not  hemorrhagic.  Spleen 
hyperplastic  and  bound  to  the  cecum  by  dense  adhesions.  Liver  yellowish. 
Kidneys  pale,  with  a  few  minute  petechia3.  Mesenteric  and  gastric  glands 
enlarged  and  hemorrhagic.  Small  intestines  little  changed.  Colon  and 
cecum  show  hemorrhagic  areas  on  their  serous  surface.  Mucosa  thickened 
and  covered  throughout  with  a  diphtheritic  layer  yellowish  in  places,  black 
in  others.  Button  ulcers  show  above  this  membrane.  Lungs  slightly  con- 
gested. Tracheal  glands  hemorrhagic.  Pure  cultures  of  bacillus  pyocyaneus 
were  obtained  from  the  spleen,  liver,  kidney,  and  mesenteric  glands. 

SECOND    PASSAGE,       FIEST    FILTRATE. 

The  blood  of  this  hog  (No.  114)  was  mixed  with  two  parts  of  sterile 
physiological  salt  solution  and  filtered  through  a  large 'Berkefeid  filter  (Labo- 
ratory Cylinder  No.  2),  using  suction.  The  filtrate  was  removed  from  the 
sterile  suction  flask  by  means  of  a  sterile  pipette,  filled  into  a  sterile  syringe, 
sealed,  immersed  in  five  per  cent  carbolic  acid  solution,  and  wrapped  in 
cotton  moistened  with  the  same  disinfectant.  So  wrapped,  the  syringe  was 
handed  to  a  second  person,  who  had  never  been  near  the  diseased  hogs,  nor 
in  the  room  where  the  filtration  took  place.  This  person  made  the  injection. 
The  animal  chosen  was  a  Chester-white  pig,  No.  117,  weighing  25  pounds.  It 
eceived  (June  17)  intraperitoneally  10  c.c.  of  the  above  filtrate,  equivalent  to 


362  McClintock,  Boxmeyee,  and  Siffee 

3.3  c.c.  of  undiluted  serum.  The  animal  was  placed  in  an  isolated  pen  at  a 
distance  from  the  diseased  hogs.  The  stableman  who  attended  him  never 
came  in  contact  with  diseased  hogs  nor  their  attendant.  The  same  attendant 
cared  for  some  uninoculated  hogs  during  this  time,  which  remained  well 
during  the  experiment,  but  later  succumbed  to  the  disease  after  a  normal 
incubation  period  when  exposed  to  infection.  The  animal  remained  active, 
but  lost  weight,  and  became  much  emaciated.  A  few  days  before  death,  he 
showed  discolored  skin,  tottering  gait,  and  blood-stained  urine.  He  died 
during  the  night  of  July  11,  21  days  after  inoculation. 

Postmortem:  Body  emaciated.  Skin  discolored  and  scabby.  Inguinal 
and  cervical  lymph-glands  swollen  and  intensely  hemorrhagic.  Numerous 
small  subcutaneous  hemorrhages.  Peritoneal  cavity  contains  considerable 
bloody  serous  exudate,  in  which  are  particles  of  fibrin.  The  bladder  has  the 
appearance  of  a  large  blood-clot;  the  walls  are  greatly  thickened  and  infil- 
trated with  blood.  Both  large  and  small  intestines  show  numerous  hemor- 
rhagic spots  on  the  serous  surface.  The  mucosa  of  colon  and  cecum  show 
numerous  large  yellowish  button  ulcers  with  black  centers.  These  ulcers  are 
occasionally  confluent.  The  lungs  show  numerous  petechiae  and  a  nodule  of 
walnut  size.  Heart  pale  and  flabby.  Visceral  pericardium  dotted  with  many 
small  ecchymotic  spots. 

Portions  of  the  filtrate  with  which  the  above  hog  was  injected  were 
tested,  as  follows : 

Five  c.c.  was  added  to  500  c.c.  of  broth. 

Two  c.c.  was  added  to  500  c.c.  of  broth  covered  with  oil. 

The  first  flask  showed,  after  some  days,  a  slight  cloudiness.  Subcultures 
and  injection  of  2.5  c.c.  of  the  flask  contents  subcutaneously  into  a  rabbit 
failed  to  show  any  bacteria.  Microscopic  examination  showed  numerous 
minute  granules  adhering  together  in  clumps.  These  were  difficult  to  stain 
with  the  ordinary  anilin  dyes,  and  probably  were  precipitated  albuminous 
substances. 

A  guinea  pig  (300  grams)  received  seven  c.c.  subcutaneously  without 
efl'ect. 

A  mouse  (20  grams  weight)  received  two  c.c.  subcutaneously.     No  effect. 

Eabbit  No.  117,  five  c.c.  subcutaneously,  without  effect. 

Rabbit  No.  116,  five  c.c.  intravenously.  Died  on  the  39th  day  from  pneu- 
monia. The  animal  showed  signs  of  snuffles  before  death.  Bipolar  staining 
bacilli  were  found  in  large  numbers  in  the  pus  from  pleural  exudate. 

THIRD    PASSAGE,    SECOND    FILTRATE. 

The  blood  and  peritoneal  fluid  of  hog  No.  117  were  left  in  the  refrigerator 
for  11  days,  then  diluted  with  three  parts  of  sterile  salt  solution,  filtered  first 
through  paper,  then  passed  twice  through  a  large  Berkefeld  filter  and  finally 
through  a  Chamberland  Pasteur  F  filter  using  suction.  This  filtrate  was  filled 
into  a  syringe,  sealed  and  handled  under  the  same  precautions  as  the  first 
filtrate.  Cultures  on  agar  and  broth  gave  no  signs  of  growth.  Rabbit  No. 
121  received  subcutaneously  nine  c.c.  of  this  filtrate  without  result.  Rabbit 
No.  120,  five  c.c.  intravenously  without  effect. 


Studies  on  Hog  Cholera  363 

Mouse  No.  150,  (15  grains)  one  c.c.  subcutaneously.  Found  dead  on  the 
second  day.  Xo  bacilli  could  be  found  at  the  point  of  inoculation  upon  micro- 
scopic examination. 

Hog  No.  121,  Chester-white,  weighing  20  pounds,  received  subcutaneously 
20  c.c.  (equal  to  five  c.c.  serum )  of  the  filtrate  from  hog  No.  117.  This  animal 
was  kept  in  a  dog-cage  in  an  animal  room  at  a  distance  from  the  diseased 
hogs,  and  neither  the  person  who  injected  the  animal  nor  the  attendant  ever 
came  in  contact  with  any  diseased  swine,  nor  into  the  room  where  infectious 
material  was  handled.  No  one  that  had  handled  infected  material  entered 
the  room  during  the  period  of  isolation.  The  animal  showed  on  the  11th  day 
distinct  signs  of  disease,  reddening  of  the  skin,  loss  of  appetite,  and  intense 
thirst.  On  the  10th  day  a  marked  diarrhea  set  in  and  the  pig  lay  crouched  in 
the  corner  of  the  cage  from  which  it  refused  to  move.  On  the  17th  day  after 
inoculation,  the  animal,  being  apparently  moribund,  was  chloroformed  and 
bled  from  the  carotid  artery,  the  blood  being  collected  aseptically. 

Postmortem:  Much  emaciated.  Skin  discolored.  Ears  purplish.  Ingui- 
nal glands  greatly  swollen  with  some  hemorrhagic  lobules.  Spleen  normal, 
liver  normal.  Cortex  of  kidneys  pale  with  a  few  petechiae.  Mesenteric  and 
retroperitoneal  glands  swollen  and  slightly  hemorrhagic.  Pancreas  normal. 
Colic  and  cecal  mucosa  free  from  ulcers,  but  dotted  with  exceedingly  numer- 
ous minute  hemorrhages.  Lungs  with  small  sub-pleural  hemorrhages.  Post- 
tracheal  glands  hemorrhagic  and  edematous.  Blood  smears  show  a  consider- 
able number  of  nucleated  red  corpuscles. 

Cultures  were  made  from  the  spleen,  liver,  kidneys  and  lymph  glands. 
Broth  tubes  from  the  first  two  showed  a  few  large  cocci.  The  others  were 
sterile. 

The  blood  serum  of  this  animal  failed  to  agglutinate  in  20  hours  either 
the  Pontiac  swine  dysentery  or  a  typical  hog  cholera  bacillus,  in  dilutions  of 
1-100.  Cultures  and  inoculations  of  rabbits  made  with  the  blood  serum  of 
this  hog  showed  no  bacteria. 

FOUETH  PASSAGE.   UNFILTEEED  SEEUM. 

Since  the  first  two  passages  and  flltrations  should  have  eliminated  all 
filterable  organisms,  and  as  the  severity  of  the  lesions  was  apparently  decreas- 
ing it  was  decided  to  return  to  unflltered  serum  in  hopes  that  the  more  char- 
acteristic lesions  of  the  disease  would  reappear. 

Hog  No.  134,  Berkshire,  weight  20  pounds.  Received  subcutaneously  five 
c.c.  of  the  unflltered  sterile  serum  of  hog  No.  124.  The  same  .precautions  in 
regard  to  the  syringe  and  the  isolation  of  the  animal  wqre  observed  as  before. 
On  August  26,  the  1.3th  day  after  inoculation  plain  signs  of  the  disease  were 
present.     The  animal  died  September  6,  24  days  after  inosculation. 

Postmortem:  Slight  loss  of  flesh.  Redness  of  skin  of  abdomen  and  chest. 
Inguinal  lymph  glands  enlarged  and  hemorrhagic.  Considerable  quantity  of 
fluid  exudate  in  the  peritoneal  cavity.  Intense  fibrinous  peritonitis,  thick 
layers  of  fibrinous  exudate  matting  together  the  viscera  and  covering  liver 
and  spleen.  The  whole  length  of  small  intestines  hemorrhagic,  mucosa 
hyperemic.     Cecum  and  adjoining  portions  of  colon  show  numerous  round 


364  McClintock,  Boxmeyer,  and  Siffee 

depressed  ulcers  from  three  to  eight  mm.  in  diameter.  A  circular  ulcerated 
ring  around  the  ileocecal  valve.  The  remaining  portion  of  the  large  intes- 
tines shows  scattered  hemorrhagic  areas.  Spleen  slightly  enlarged.  Liver 
and  kidneys  apparently  normal.  Fibrinous  pleuritis  and  pericarditis.  Parie- 
tal pericardium  adherent  to  the  heart.  Small  congested  and  edematous 
areas  in  the  apices  of  the  lungs,  other  portions  normal.  Posttracheal  glands 
enlarged  and  hemorrhagic. 

Cultures  made  from  liver,  spleen  and  kidneys  all  showed  growth.  None 
of  the  organisms  obtained  were  agglutinated  either  by  a  hog  cholera  or  a 
swine  dysentery  serum. 

In  addition  to  the  above  series  a  small  white  hog  No.  116,  weight  20 
pounds,  was  inoculated  with  a  filtrate  made  by  steeping  the  viscera  brought 
from  Pontiac  in  salt  solution,  passing  through  paper  and  a  Berkefeld  filter. 
The  first  two  weeks  he  apparently  gained  in  weight,  but  later  appeared  dull, 
sniffled  and  lay  in  the  corner  of  the  cage.  He  died  on  the  35th  day  after 
inoculation.  The  precautions  against  accidental  infection  were  the  same  as 
observed  before. 

Postmortem:  Reddening  of  the  ears.  Inguinal  glands  greatly  enlarged 
with  minute  punctiform  hemorrhages.  Cervical  lymph  glands  enlarged  and 
hemorrhagic.  Spleen  dark,  not  enlarged.  Liver  normal.  Small  fibrinous 
particles  adhered  to  the  spleen  and  serus  covering  of  the  intestines.  Mesen- 
teric glands  enlarged.  Gastric  and  mesocolic  glands  hemorrhagic.  Small 
intestines  filled  with  yellow  fluid  feces.  Kidneys  pale  with  a  few  hemorrha- 
gic spots.  Large  intestines  practically  normal  except  at  the  base  of  the  ileo- 
cecal valve  where  were  found  two  small  yellowish  ulcers.  Apices  of  both 
lungs  hepatized.  Posttracheal  lymph  glands  edematous  and  hemorrhagic. 
Considerable  blood-stained  purulent  exudate.  The  Pontiac  bacillus  was 
isolated  from  the  mesenteric  glands  of  this  hog. 

Early  in  the  winter  some  preliminary  experiments  had  been 
made  with  infectious  material  fi-om  the  outbreak  that  occurred  at 
Hastings,  Michigan,  in  September,  1903.* 

Neither  in  material  from  this  epizootic  brought  from  the  field  nor  in  the 
animals  infected  in  the  laboratory  could  hog  cholera  bacilli  ever  be  isolated, 
nor  did  the  blood  serum  of  any  of  these  animals  exhibit  any  agglutinating 
power  for  hog  cholera  bacilli,  in  excess  of  that  normally  present. 

The  liver  and  spleen  of  hog  No.  47  (an  acute  hemorrhagic  case  produced 
by  feeding  viscera  brought  from  Hastings)  were  steeped  in  salt  solution  and 
the  fluid  filtered  first  through  paper,  then  through  a  No.  6  Berkefeld  filter. 

*This  epizootic  was  complicated  in  one  herd  by  foot  lesions  from  which  an  anerobic 
bacillus,  a  member  of  the  malignant  edema  group,  was  isolated.  The  disease  resembled 
blackleg  in  being  usually  confined  to  one-quarter  of  the  body.  The  lesions  consisted  of  a 
hemorrhagic  edema  with  emphysema  extending  along  the  limb  into  the  foot.  Ulcers  were 
present  on  the  foot  and  between  the  toes.  In  one  animal  the  entire  liver  was  emphysema- 
tous, and  had  the  appearance  and  feel  of  luug  tissue.  A  rabbit  inoculated  with  edema  fluid 
from  this  animal  showed  a  similar  condition  of  the  liver.  These  lesions  were  reproduced  in 
the  laboratory  in  hogs  fed  infected  carcasses.  The  bacillus  died  out  before  it  could  be  more 
carefully  studied. 


Studies  on  Hog  Cholera  365 

Hog  No.  62,  a  Chester-white,  weighing  30  pounds,  received  intraperitoneally 
four  c.c.  of  the  above  filtrate.  No  bacteria  were  detectable  in  this  filtrate  by 
cultures  or  animal  inoculation.  Fifteen  days  after  inoculation,  the  pig  showed 
diarrhea  and  blood-stained  feces.  Plates  (Drigalski  &  Conradi)  made  from 
the  feces  failed  to  reveal  hog  cholera  bacilli.  Death  occurred  in  38  days. 
Autopsy  showed  discoloration  of  the  skin,  hemorrhagic  lymph  glands,  small 
dark  spleen,  ecchymotic  mucous  and  serous  membranes,  and  the  colon  and 
cecum  had  scattered  thickly  throughout  their  length  small  yellowish  nodules 
about  the  size  of  a  goose  shot.  These  nodules  were  due  to  the  distention  of 
Klein's  glands  by  cheesy  pus.  Cultures  showed  a  variety  of  bacilli  but  no 
hog  cholera  bacilli  could  be  isolated. 

Hog  No.  58,  Chester-white,  30  pounds,  received  intraperitoneally  two  c.c. 
of  the  edema  fluid  of  hog  No.  47,  filtered  through  a  small  Berkefeld  filter. 
Cultures  and  animal  inoculations  of  the  filtrate  showed  no  bacilli.  The 
course  of  the  disease  was  similar  to  that  of  hog  No.  62.  The  animal  died  on 
the  SSlh  day  after  inoculation.  The  pathological  appearances  were  similar 
to  those  in  hog  No.  62.  The  colon  and  cecum  showed  numerous  raised  button 
ulcers  6  to  20  mm.  in  diameter  scattered  along  their  length,  and  a  large  necro- 
tic patch  at  the  ileocecal  valve. 

Hogs  No.  109  and  No.  110,  weighing  25  pounds  each,  received  subcu- 
taneously  respectively  14  c.c.  of  a  Berkefeld  No.  2  and  10  c.c.  of  a  Pasteur  P. 
filtrate  of  an  emulsion  of  the  viscera  of  hog  No.  106  (infected  with  material 
from  hog  No,  47).  Hog  No.  109  died  on  the  20th  and  hog  No.  110  on  the  24th 
day  a'ter  infection.  No  characteristic  lesions  were  present.  Both  showed 
slight  pneumonia  and  hyperemia  of  the  intestinal  mucosa.  Cultures  from 
hog  No.  110  gave  no  growth.  Hog  No.  109  gave  pure  cultures  of  streptococci. 
The  blood  of  neither  hog  agglutinated  hog  cholera  bacilli  in  dilutions  of 
1-.300. 

In  summarizing  the  data  we  find  that  of  five  hogs  inocu- 
lated with  sterile  Berkefeld  filtrates  derived  from  two  distinct 
outbreaks,  all  died  within  a  period  varying  from  19  to  38  days,  a 
period  which  coincides  with  that  observed  in  cases  of  natural 
infections  and  those  produced  by  feeding  infected  material.  Four 
of  these  (Nos.  62,  58,  114,  and  116)  showed  the  characteristic 
intestinal  ulcers  of  hog  cholera.  Both  hogs  (Nos.  110  and  124) 
inoculated  with  Pasteur  filtrates  died,  one  on  the  24th,  the  other 
on  the  19th  day  after  inoculation.  Though  the  source  of  the 
infecting  filtrate  was  different  (one  Hastings,  the  other  Pontiac), 
both  showed  similar  lesions,  which,  while  not  characteristic  of 
hog  cholera  are  frequently  found  in  natural  infections.  The 
blood  serum  of  one  of  these  (hog  124),  when  injected  into  another 
hog  (No,  134),  produced  the  characteristic  lesions  of  hog  cholera, 
the  animal  dying  in  the  usual  period,  namely,  24  days. 


366  McClintock,  Boxmeyee,  and  Siffee 

The  above  observations  are  in  agreement,  on  the  whole,  with 
those  of  the  Bureau  of  Animal  Industry,  but  differ  in  that  we 
had  to  deal  with  the  chronic  form  of  the  disease,  while  their  virus 
always  gave  rise  to  the  acute.  In  an  attempt  to  produce  an  acute 
infection  a  100-pound  hog  received  an  injection  of  five  c.c.  of 
virulent  blood  serum  into  the  ear  vein.  He  showed  on  the  eighth 
day  a  sharp  rise  of  temperature  and  refused  food.  On  the  14th 
day  he  had  diarrhea  and  other  symptoms  of  the  disease,  but 
was  still  strong  and  death  would  probably  not  have  occurred  for 
a  couple  of  weeks.  As  virulent  blood  was  required  for  immuniz- 
ing experiments,  this  animal  was  bled  to  death  on  the  15th  day 
after  inoculation.  This  experiment  may  be  considered  a  failure 
so  far  as  the  production  of  an  acute  form  of  hog  cholera  is  con- 
cerned. The  same  procedure  applied  to  smaller  hogs  would 
probably  have  been  successful,  as  they  are  much  more  susceptible 
to  infection. 

The  failure  to  isolate  from  the  Hastings  outbreak  a  hog 
cholera  bacillus,  although  a  number  of  animals  were  examined 
and  the  infection  passed  from  animal  to  animal  for  several  genera- 
tions in  the  laboratory,  the  ease  with  which  hog  cholera  bacilli 
can  be  recovered  when  once  introduced  into  the  system,  the 
difficulty  of  infecting  swine  with  any  but  large  quantities  of  the 
most  virulent  bacilli,  the  readiness  with  which  the  disease  is 
naturally  spread,  taken  together  with  the  results  from  the  above 
filtration  experiments  seem  to  show  conclusively  that  the  hog 
cholera  bacillus  is  not  the  primary  factor  in  the  production  of  hog 
cholera,  if  the  disease  studied  were  such. 

Yet  let  us  assume  that  the  hog  cholera  bacillus  produces  in 
the  blood  of  the  infected  animals  a  toxin,  and  that  none  of  the 
toxin  is  used  up  in  producing  the  disease.  In  its  serial  passage 
through  hogs  No.  117  and  No.  121,  calculating  the  blood  as  five 
per  cent  of  the  body  weight,  we  would  have  diluted  the  original 
toxin  20,000  times.  The  third  hog  received  five  c.c.  of  this 
dilution  or  .00025  c.c.  of  the  original,  which  is  equivalent  to 
,000000025  c.c.  per  gram  weight  of  hog — a  toxin  whose  potency 
has  never  been  equalled  except  possibly  in  the  case  of  purified 
tetanus  toxin.     Up  to  the  present  no  substance  of  any  great  toxic 


Studies  on  Hog  Choleea  367 

power  has  been  obtained  from  bacilli  of  the  hog  cholera  typhoid 
group,  and  toxins  of  the  diphtheria  tetanus  type  are  wholly 
unknown  among  its  members.  In  the  present  state  of  our  knowl- 
edge, it  seems  unreasonable  to  suppose  that  a  toxin  of  so  great  a 
potency  is  formed.  The  following  experiment  furnishes  addi- 
tional evidence  that  the  disease-producing  agent  is  not  an 
unorganized  toxin.  Hog  No.  125,  weighing  20  pounds,  received 
fivec.c.  of  chloretone-saturated  blood  of  hog  No.  115.  During  the 
20  days  he  was  under  observation  he  showed  no  signs  of  disease, 
and,  when  exposed  later  with  an  infected  animal,  died  showing  the 
typical  lesions  of  hog  cholera.  Two-thirds  of  a  cubic  centimeter 
of  the  same  blood  without  the  addition  of  chloretone  produced 
typical  hog  cholera  when  injected  subcutaneously  into  hogs  of 
the  same  size.  Tri.-  chlor.-  tertiary-  butyl-alcohol  has  been  shown 
in  this  laboratory  to  be  without  effect  upon  diphtheria  and  tetanus 
toxins,  but  rapidly  destructive  to  non-spore-bearing  organisms. 
This  experiment,  in  connection  with  the  preceding,  would  seem 
to  show  that  the  infecting  agent  was  some  organism  readily  killed 
by  mildly  germicidal  substances,  and  capable  of  passing  through 
the  pores  of  Berkefeld  and  Chamberland  filters. 

The  above  facts  appear  to  place  hog  cholera  in  the  group  of 
diseases  among  which  are  rinderpest,  peripneumonia,^  and  foot  and 
mouth  disease,"^  all  of  which  are  capable  of  being  transmitted  by  a 
filterable  virus.  With  the  exception  of  peripneumonia,  the  causative 
organism  of  none  of  these  has  ever  been  cultivated  or  seen  under 
the  microscope.  The  size  of  the  organism  of  peripneumonia 
which  has  been  cultivated  by  Nocard  and  Roux  lies  at  the  limit 
of  microscopic  vision.  The  pathological  lesions  found  in  hog 
cholera  are  quite  similar  to  those  described  for  rinderpest,  as  seen 
in  the  Philippines,^  and  the  disease  is  probably  more  closely 
related  to  this  than  to  any  of  the  others. 

The  virus  differs  from  that  of  rinderpest,  however,  in  the 
greater  readiness  with  which  it  passes  through  filters*  and  its 

1  NocAED,  Handbuch  d.  Path.  Microorganismen,  1903,  3,  p.  682. 

2  LOFFLEE,  Centralbl.  f.  Bakt.,  1898,  23,  Abth.  1,  p.  371. 

3  JoBLiNG,  Report  of  the  Philippine  Government  Laboratories,  1903,  p.  363. 
*  NicoLLE  and  Adil-Bey,  Ann.  de  VInst.  Pasteur,  1902, 16,  p.  56. 


368  McClintock,  Boxmeyer,  and  Siffee 

greater  resistance  to  physical  and  chemical  agents.  Koch^  states 
that  the  virus  of  rinderpest  dried  at  31°  C.  was  inert;  also  that 
the  addition  of  10  per  cent  glycerin  to  virulent  blood  destroyed 
its  infectious  qualities. 

I  have  found  that  the  blood  serum  of  diseased  hogs  dried  at 
37°  C.  or  the  serum  to  which  one-third  its  bulk  of  glycerin  had 
been  added  did  not  suffer  in  virulence.  When  tested  one  month 
after  preparation,  it  caused  apparently  as  severe  a  form  of  disease 
as  the  fresh  serum. 


IV. 
IMMUNIZATION  AGAINST  HOG  CHOLERA. 

Charles  H.  Boxmeyer  and  Charles  T.  McClintock. 

When  preliminary  experiments  had  shown  that  hog  cholera 
could  be  transmitted  by  a  filterable  virus,  the  method  employed 
by  Kolle  and  Turner"  in  combating  rinderpest  at  Kimberly, 
South  Africa,  suggested  itself  as  applicable  in  this  disease  also. 
This  method  consisted  in  the  simultaneous  injection  of  virulent 
blood  and  the  blood  serum  of  an  immune  animal.  The  source  of 
the  immune  blood  was  at  first  cattle  that  had  recovered  from  the 
disease  either  naturally  contracted  or  produced  by  inoculation 
with  the  blood  of  diseased  animals.  Later  the  serum  of  animals, 
subsequently  hyper-immunized  by  the  injection  of  large  quanti- 
ties of  virulent  blood,  was  used  with  still  better  success.  Roger's^ 
modification,  the  injection  of  a  second  somewhat  increased 
quantity  of  infectious  blood  after  the  disappearance  of  the 
reaction  from  the  first  injection  of  virulent  and  immune  blood, 
also  seemed  worthy  of  trial.  In  addition,  attempts  were  made  to 
immunize  hogs  by  the  injection  of  diseased  blood  dried  at  a 
temperature  of  37°  C.  and  blood  to  which  one-third  of  its  weight 
of  glycerin  was  added. 

EXPERIMENT  WITH  DRIED  BLOOD. 

Hog  120,  Chester-white  female,  20  pounds,  received  subcutaneously,  dis- 
solved in  normal  salt  solution,  0.1  gram  of  diseased  blood  which  had  been 

1  Dent.  med.  Wchmchr.,  1897,  23,  pp.  225-41.  2  Deut.  med.  Wchnschr.,  1897,  pp,  793,  818 

3  Zeitschr.  f.  Hyg.  ti.  Infectionsk.,  1900,  35,  p.  59. 


Studies  on  Hog  Cholera 


369 


dried  at  37°  C.  for  36  hours,  14  days  previously.  On  the  third  day  the  animal 
showed  loss  of  appetite  wdth  an  inclination  to  remain  in  its  litter.  On  the  fifth 
day  it  refused  food.  The  skin  showed  a  bright  flush,  and  a  marked  diarrhea 
was  present.  There  was  but  a  slight  rise  in  temperature  (see  tempera- 
ture curve).  The  animal  gradually  grew  worse  and  died  on  the  29th  day 
after  inoculation.  Autopsy  showed  typical  hog  cholera  lesions,  the  colon  and 
cecum  containing  numerous  ulcers  both  of  the  deep  and  button  type  scattered 
along  their  length. 

HOG  1.0 
Days    12        .3        1        5       6        7        8        9       10      11      12      13      14       15      16      17      18      19 


f.         ^^l^r-                                 A 

,or      \    ,,       ^^X    ^^-/    ^2    ^"^-    ,       j\          ^-' 

X'^X^       V                                      ^^U        ^       4 

,03.           ^       ^             '^                                             ^              V^^ 

A 
-Vaccinated 


Afebrile  type  of  hog  cholera. 


EXPEEIMENTS  WITH  GLYCERINATED  BLOOD. 

I.  Hog  121,  male  Chester-white,  weighing  20  pounds,  received  subcutane- 
ously  one  c.c.  of  freshly  prepared  glycerinated  blood  (blood  two  parts,  glycerin 
one  part).  On  the  third  day  it  showed  loss  of  appetite  and  languor.  On  the 
fifth  day  refused  food  and  was  constipated.  On  the  seventh  day  the  tempera- 
ture had  risen  to  105  F.,  and  attained  a  maximum  of  107°  F.  on  the  12th  day- 
The  animal  being  moribund  was  chloroformed  on  the  20th  day.  Autopsy 
showed  typical  lesions,  colon  and  cecum  being  covered  with  numerous 
depressed  circular  ulcers.     The  rectal  mucosa  was  also  badly  ulcerated. 


Days 

108° 


107° 
103" 
105° 
104° 
103° 


HOG  121 
1        2        3        4        5       6        7       8        9      10      11      12      13      14      15      16     17     18     19     20 

^^"V 

I    V      ^^'^^ 

t     ^:^  /   r    r  " 

f         ±        ^^  _  a:3:4~  " 

t\         A     fA                                             —%    V^ 

^     -~\4'J^l                         _     ^_V  L 

^^          -.\-A     ^X 

^^\,u^j    i 

21 


A 


1  CO.  Glyc.  Blood 


A 
20  CO.  immune  Serum 

Febrile  type  of  hog  cholera. 


A 

Dying  chloroformed 


II.  Hog  129,  Chester-white  female,  weighing  40  pounds,  received  one  c.c. 
of  same  glycerinated  blood  now  24  days  old.  It  showed  loss  of  appetite  on 
the  third  day,  but  still  fed  a  little.  On  the  15th  day  the  temperature  rose 
suddenly,  animal  showed  reddening  of  the  skin,  refused  to  eat  and  appeared 
very  ill.  Death  occurred  on  the  31st  day  after  injection.  Autopsy  showed 
typical  hog  cholera  lesions. 


370 


McClintock,  Boxmeyer,  and  Siffek 


These  experiments  showed  no  attenuation  of  the  virus. 

Hog  125  received  five  c.c.  of  the  same  diseased  blood  saturated  with 
chloretone.  During  the  20  days  it  was  under  observation,  the  animal  showed 
no  signs  of  disease,  and,  when  exposed  with  a  diseased  hog,  contracted  the 
disease  after  the  regular  incubation  period,  and  died  on  the  22d  day  after 
exposure,  showing  numerous  ulcers  in  the  colon  and  cecuiii.  In  this  instance 
the  chloretone  had  apparently  destroyed  the  infecting  agent  and  the  injection 
produced  no  immunity. 

EXPERIMENTS    WITH    SIMULTANEOUS    INJECTION    OF    VIRULENT     AND 

IMMUNE    BLOOD, 

Two  hogs  furnished  the  immune  serum  employed.  One  of  these,  hog  No. 
113,  was  repeatedly  fed  infected  viscera.  These  failed  to  produce  any  notice- 
able disease.  The  animal  was  subsequently  further  immunized  by  the  injec 
tion  of  considerable  quantities  of  filtrates.  He  was  chloroformed  and  bled 
from  the  carotid.  Postmortem  showed  no  marked  lesions.  The  colon  and 
cecum  were  free  from  ulcers  or  scars. 

The  second  hog,  No.  108,*  was  also  fed  infected  viscera  without  effect  and 
later  confined  with  a  diseased  animal,  from  which  he  contracted  the  disease 
which  failed  to  prove  fatal,  but  left  him  badly  stunted.  He  was  further 
immunized  by  the  feeding  and  injection  of  large  quantities  of  infectious 
material  which  in  no  instance  produced  any  further  I'eaction.  He  was  chloro- 
formed and  bled  from  the  carotid. 

In  both  cases  the  blood  serum  was  preserved  by  the  addition  of  four 
parts  per  1000  of  trikresol. 

Hog  No.  118,  Chester-white,  weight  30  pounds,  received  subcutaneously, 
on  one  side,  one  c.c.  of  diseased  blood,  and  on  the  other  side,  five  c.c.  of 
immune  blood.  The  animal  showed  a  rise  in  temperature  on  the  fourth  day 
with  loss  of  appetite  and  symptoms  of  hog  cholera.  The  temperature  attained 
its  maximum  on  the  sixth  day,  and  fell  to  normal  on  the  seventh,  where  it 
remained  till  the  11th  day,  on  which  a  relapse  set  in,  and  the  animal  died  on 
the  34th  day. 

HOG -118. 


Days      1       2 
10.-r 

3       4        -5 

6 

7        8        9       10     11      12      13      14 

-          ^      ^- 

\ 

104^                  5 

^^ 

\.                            -          rt 

U  V 

1       ~      ~ 

\                                        '    -      ^ 

103               ^ 

V'^ 

V  ^     -  X-V^ 

^A   A        i-        ^- 

102' 

\/^  ^J- 

^  g  ^ 

A 
Virulent  and  immune  blood 

*This  hog  was  fed  large  quantities  of  the  viscera  of  two  hogs  that  had  been  killed  by 
intravenous  injection  of  the  Arkansas  hog  cholera  bacillus.  He  failed  to  show  any  effects 
from  this  feeding  though  under  observation  six  weeks.  Later  he  was  fed  viscera  from  hogs 
naturally  infected.  These,  however,  had  been  kept  for  some  months  in  the  refrigerator,  and 
had  probably  largely  lost  their  infectious  qualities. 


Studies  on  Hog  Choleea 


371 


Compare  with  the  following  : 


HOG  119. 


Days  1 
103  = 

104° 

103° 


10      II      12      13      11 


102= 


-v"^^ 

5          ^^  ^^'^   - 

L-Ok.                      jL                                ^       ^                  ^ 

^\  f^/              ^'S     r-^^  ^-s 

:  All  ^                 XJ          -V'^ 

A 


Diseased  Blood  only 

Hogs  Nos.  126,  127  and  131,  each  weighing  20 
pounds,  received  in  the  same  manner  as  the  preceding 
one  c.c.  each  of  diseased  blood  (No.  126  dried,  No.  127 
and  No.  131  glycerinated  blood)  and  10  c.c.  of  immune 
blood.  All  showed  loss  of  appetite  and  slight  rise  of 
temperature  on  the  fourth  day.  The  temperature 
attained  its  maximum,  106  to  107°  F.,  on  the  seventh 
or  eighth  day,  and  in  the  case  of  126  and  127  it  returned 
to  normal  on  the  11th  day,  and  showed  no  further  rise. 
In  the  case  of  No.  131  the  temperature  remained  high 
until  the  13th  day,  after  which  it  dropped  and  remained 
normal. 

On  the  16th  day  hogs  No.  126  and  No.  127  were  con- 
fined with  a  diseased  hog.  On  the  sixth  day  after 
exposure  their  temperature  rose  suddenly  and  a  mild 
form  of  hog  cholera  set  in.  Both  animals  recovered  ; 
No.  127  without  showing  any  appreciable  loss  of  weight. 
In  the  case  of  No.  126,  however,  the  disease  continued 
longer  and  left  the  animal  somewhat  emaciated. 

Hog  No.  131,  although  constantly  exposed  with 
diseased  animals  has  shown  no  further  rise  in  temper- 
ature or  signs  of  the  disease. 

Hog  No.  130,  black  male,  weighing  10  pounds, 
received  subcutaneously  one  c.c.  of  glycerinated  dis- 
eased blood  and  10  c.c.  of  immune  blood.'  It  was 
noticed  at  the  time  of  injection  that  the  hog  had  a 
slight  cough,  some  diflBculty  in  breathing,  and  that 
there  was  a  purulent  discharge  from  the  nose.  He 
appeared  quite  sick  on  the  sixth  day,  gradually  grew 
worse,  and  died  on  the  18th  day  after  inoculation. 
Autopsy  showed  typical  hog  cholera  lesions.  In  addi- 
tion the  lungs  were  almost  completely  hepatized. 
This  was  apparently  a  case  of  latent  swine  plague 
complicating  the  vaccination  and  precipitating  the 
fatal  result. 


<  ° 


372 


McClintock,  Boxmeyee,  and  Sifpee 


Hogs  No.  122  and  123,  Chester-white  sows,  weighing  200  pounds  each, 
(sow  No.  122  was  pregnant).  Each  received  20  c.c.  of  immune  serum  subcuta- 
neously.  Two  days  afterwards  1.5  c.c.  of  glycerinated  blood.  Five  days  after- 
ward both  showed  loss  of  appetite,  slight  reddening  of  the  skin  and  indispo- 

HOG  127. 
Days  1  2  3  4  5  6  7  8  9  10  11  12  13  14  15  16  17  18  19  20  21  22  23  24  25  26  27  28  29  30  31  .32  .33  34  35  36  37  38  39  40  41  42  43  44  45 


108= 
105' 


103= 


A 


t^' 


b'^^ 


^5: 


•=7- 


llUMUl-H 


m 


-V 


ffl 


I  I  I  M 


A 
Vaccinated 


A 
Exposed 

sition.  On  the  10th  day  they  fed  normally  again.  Twenty-one  days  after 
inoculation  each  received  five  c  c.  of  virulent  blood.  Having  shown  no  effect 
from  this  injection  28  days  afterward  they  received  100  c.c.  of  virulent  blood 
and  three  and  eight  days  later  a  second  and  third  injection  of  100  c.c.  each, 
from  which  no  effect  other  than  a  slight  local  reaction  was  observed.  No. 
122  gave  birth  at  term  to  a  litter  of  healthy  pigs. 

Hogs  No.  1.32  and  No.  1.33,  Chester-whites,  weighing  100  pounds,  received 
each,  subcutaneously,  1.5  c.c.  of  glycerinated  blood  and  10  c.c.  of  immune  serum. 
They  became  sick  on  the  12th  and  14th  day,  refusing  to  eat  and  showing 
indisposition  and  a  considerable  rise  of  temperature.  On  the  20th  day  the 
temperature  fell  to  normal,  their  appetite  improved  and  they  appeared  on  the 
29th  day,  perfectly  healthy.  They  and  the  sows  Nos.  122  and  123  were  ex- 
posed to  infection  from  hogs  in  a  neighboring  pen  during  this  entire  period. 

The  results  from  this  simultaneous  method  of  immunization 
are  on  the  whole  satisfactory.  The  larger  animals  have  shown 
themselves  readily  immunized  against  the  disease,  the  process  of 
immunization  producing  no  marked  illness,  no  considerable  loss 
of  weight,  nor  tendency  to  stunt  the  growth,  items  of  the  greatest 
importance  in  the  practical  application  of  any  method.  Younger 
hogs  react  much  more  strongly  to  the  vaccination,  still  we  have 
been  successful  in  immunizing  them  also.  The  failures  can  be 
laid  in  the  case  of  No.  118  to  the  administration  of  an  insufficient 
amount  of  immune  serum,  and  in  the  case  of  No.  130  to  the 
presence  of  latent  swine  plague. 

The  secoiifl  rise  in  temperature  in  hogs  Nos.  12(3  and  127  may 
possibly  be  a  relapse,  but  more  likely  a  re-infection,  the  reaction 
from  the  vaccination  being  in  both  cases  extremely  mild  and 
probably  insufficient  to  produce  complete  immunity.      This  w^ould 


Studies  on  Hoa  Choleea 


373 


indicate  the  necessity  of  establishing  a  very  nice  balance  between 
the  amounts  of  immune  and  virulent  blood  injected  in  order  to 
0|btain  the  most  perfect  results. 

Durinar  the  entire  time  of  immunization  the  animals  were 
exposed  to  constant  possibility  of  infection,  which  may  in  some 
degree  have  influenced  our  results. 

THE    CUEATIVE    EFFECTS    OF    HOG    CHOLEEA    IMMUNE    SEEUM. 

Hog  No.  118  received  on  the  17th  day  after  infection  22  c.c.  of  the  immune 
serum  from  hog  No.  113.  This  injection  of  serum  had  no  influence  upon  the 
course  of  the  disease,  which  terminated  fatally. 

Hog  No.  119  received  30  c.c.  of  the  same  serum  on  the  18th  day  after 
infection.  This  produced  an  immediate  fall  of  temperature.  The  animal 
recovered;  the  value  of  the  serum  injection  was  not  apparent. 

Hog  No.  121  received  on  the  sixth  day  after  infection  20  c.c.  of  immune 
serum  which  produced  no  noticeable  effect  either  upon  the  temperature  or 
upon  the  course  of  the  disease  which  ended  fatally. 

Hog  No.  125  received  30  c.c.  on  the  eighth  day  with  no  effect  upon  the 
temperature  or  the  course  of  the  disease. 

The  administration  of  immune  serum  after  the  sixth  day  in  the  amounts 
given  has  been  wholly  without  result.  What  the  effect  would  be  when  given 
in  larger  quantities  and  earlier  in  the  disease,  we  have  so  far  made  no  experi- 
ments to  determine. 

The  only  record  of  any  previous  attempt  to  immunize  hogs  by 
a  similar  method  to  be  found  was  that  of  Preiz,^  who  by  the 
injection  of  the  serum  of  a  hog,  that  had  recovered  from  a  severe 

HOG  131. 
Days  1  2  3  4  5  6  7  8  9  10  11  12  13  11  15  16  17  18  19  20  21  22  23  24  25  26  27  28  29  30  31  32  33  34  35 


attack  of  hog  cholera,  into  swine  already  exposed  to  infection, 
succeeded  in  greatly  reducing  the  mortality  among  them.  We 
were  unable  to  discover  any  further  application  of  this  idea  either 
by  Preiz  or  others. 


^Ztschr.f.  Thiermedicin,  1898,2,  p.  1. 


374  McClintock,  Boxmeyer,  and  Siffee 

SUMMAEY 

1.  Hogs  may  be  immunized  against  hog  cholera  by  the  simul- 
taneous injection  of  diseased  and  immune  blood,  the  larger  ani- 
mals with  great  readiness  and  apparently  without  danger,  while 
the  smaller  animals  react  more  violently  and  some  loss  is  liable  to 
occur. 

2.  Drying  at  37°  C.  or  the  addition  of  33  per  cent  glycerin 
does  not  attenuate  the  virus  sufficiently  for  immunizing  purposes. 

3.  Moderate  doses  of  immune  serum  show  no  curative  powder. 


